Exocellular peptido-polysaccharide (glycopeptide) is first released by Penicillium charlesii immediately upon the onset of NH4 ion starvation. Prior to this, galactofuranosyl-containing substances are found associated with two membrane fractions and soluble peptido-polysaccharide is found in the intracellular supernatant fluid within a short time following NH4 ion depletion of the medium. The objectives of this research are to delineate the sequence of reactions that occur in the post translational biosynthesis of a membrane-bound galactofuranosyl-containing lipo-peptido-polysaccharide, to determine the location of this substance within the cell and to determine the factors which control the synthesis and degradation of the lipo-peptido-polysaccharide. Purified mannosyltransferases will be used along with exocellular peptido-polysaccharide, intracellular peptido-polysaccharide, and the membrane-bound lipo-peptido-polysaccharide as mannosyl acceptors to investigate the biosynthesis of the mannosyl containing polysaccharide and the mannosyl-containing oligosaccharides. Vacuoles will be isolated from P. charlesii spheroplasts and the occurrence or lack of occurrence of the lipo-peptido-polysaccharide will be determined with immunochemical and enzymatic techniques. The importance of S-adenosyl-L-methionine in initiating the degradation of the membrane-bound lipo-peptido-polysaccharide will be investigated. In particular, we will determine if conversion of ethanolamine residues in the lipo-peptido-polysaccharide to N,N-dimethyl-ethanolamine occurs during the conversion of the lipo-peptido-polysaccharide to the exocellular peptido-polysaccharide. We will determine if the lipo-peptido-polysaccharide becomes more or less stable to lytic enzymes obtained from the vacuole as a result of methylating available amino groups in the lipo-peptido-polysaccharide.